S-allyl-L-cysteine Research Articles

Effect of S-Allyl-L-Cysteine on Nitric Oxide and Cadmium Processes in Rice (Oryza sativa L. sp. Zhongzao35) Seedlings

Nitric oxide (NO) is an important signaling molecule involved in regulating plant processes to cope with abiotic stress. S-allyl-L-cysteine (SAC) is known to induce NO synthesis in animals. However, it is unknown whether SAC can trigger NO biosynthesis, regulate Cd transport, or alleviate Cd stress in plants. After being sprayed with 0.2 mM SAC, rice seedlings had a NO content that was 1.8 times higher than that of the control (ctrl) group at the ninth hour, which then gradually decreased. The expressions of Cd uptake and transport genes in the roots (including OsNRAMP5, OsNRAMP1, and OsHMA2) were markedly downregulated by 27.2%, 24.8%, and 49.1%, respectively, 72 h after SAC spraying treatment. The Cd content in seedling roots’ cell wall (CW) components significantly increased by 43.5% compared to that of the ctrl group. The Cd content in the shoots and roots decreased by 49.0% and 29.8%, respectively. Cd stress in the seedlings was also substantially alleviated. In conclusion, spraying rice seedlings with SAC triggered an increase in NO synthesis, regulated the expression of genes related to Cd transport, increased Cd fixation in the root CW components, and reduced Cd accumulation in the roots and shoots.

Therapeutic effects of S-allyl-L-cysteine in a mouse endometriosis model and its immunomodulatory effects via regulation of T cell subsets and cytokine expression.

Endometriosis is a female hormone-dependent gynecological disorder characterized by chronic inflammation. Therefore, the development of novel treatment strategies that can diminish the side effects of the long-term use of hormone-based drugs has been emphasized. S-Allyl-L-cysteine (SAC) is the major constituent of aged garlic extracts. Although the therapeutic effects resulting from the antioxidant properties of SAC have been extensively studied in inflammatory diseases, the therapeutic efficacy of SAC in endometriosis has not been described. In this study, we investigated the therapeutic potential of SAC for endometriosis using a mouse model. An endometriosis mouse model was surgically induced, and oral treatment with 30mg/kg SAC was administered daily for 28 days. The development of endometriotic lesions was assessed by histological analysis, and the expression profiles of adhesion-, apoptosis-, and inflammation-related genes were evaluated by PCR. Flow cytometric analysis of mouse spleen was conducted to assess changes in lymphocyte subpopulations. SAC treatment significantly inhibited endometriotic lesion growth. Transcriptional expression analysis revealed the antiadhesion and apoptosis-promoting effects of SAC. In particular, SAC showed an effective immune modulatory response by altering splenic CD4+ and CD8+ T cell subsets and inflammatory cytokine production in the spleen and endometriotic lesions. This study newly elucidates the inhibitory effects of SAC on the growth of endometriosis in a mouse model and describes its immunomodulatory effects.

Phenylephrine Enhances the Mitogenic Effect of S-Allyl-L-cysteine on Primary Cultured Hepatocytes through Protein Kinase C-Induced B-Raf Phosphorylation.

The co-mitogenic effects of the α1-adrenoceptor agonist phenylephrine on S-allyl-L-cysteine (SAC)-induced hepatocyte proliferation were examined in primary cultures of adult rat hepatocytes. The combination of phenylephrine (10-10-10-6 M) and SAC (10-6 M) exhibited a significant dose-dependent increase in the number of hepatocyte nuclei and viable cells compared to SAC alone. This combination also increased the progression of hepatocyte nuclei into the S-phase. The potentiating effect of phenylephrine on SAC-induced cell proliferation was counteracted by prazosin (an α1-adrenergic receptor antagonist) and GF109203X (selective protein kinase C (PKC) inhibitor). In addition, PMA (direct PKC activator) potentiated the proliferative effects of SAC similarly to phenylephrine. In essence, these findings suggest that PKC activity plays a crucial role in enhancing SAC-induced cell proliferation. Moreover, the effects of phenylephrine on SAC-induced Ras activity, Raf phosphorylation, and extracellular signal-regulated kinase 2 (ERK2) phosphorylation were investigated. Phenylephrine (or PMA) in combination with SAC did not augment Ras activity, but further increased ERK2 phosphorylation and its upstream B-Raf phosphorylation. These results indicate that PKC activation, triggered by stimulating adrenergic α1 receptors, further amplifies SAC-induced cell proliferation through enhanced ERK2 phosphorylation via increased B-Raf-specific phosphorylation in primary cultured hepatocytes.

Organosulfur Compounds, S-Allyl-L-Cysteine and S-Ethyl-L-Cysteine, Target PCSK-9/LDL-R-Axis to Ameliorate Cardiovascular, Hepatic, and Metabolic Changes in High Carbohydrate and High Fat Diet-Induced Metabolic Syndrome in Rats.

Metabolic syndrome (MetS) is an ever-evolving set of diseases that poses a serious health risk in many countries worldwide. Existing evidence illustrates that individuals with MetS have a 30%-40% higher chance of acquiring type 2 diabetes mellitus (T2DM), cardiovascular disease (CVD), or both. This study was undertaken to uncover the regulatory role of natural organosulfur compounds (OSCs), S-allyl-L-cysteine (SAC), and S-ethyl-L-cysteine (SEC), in targeting high carbohydrate high fat (HCHF)-diet-induced MetS-associated risk management. Our findings suggested that SAC and SEC ameliorated HCHF-diet-induced diabetic profiles, plasma lipid and lipoprotein level, liver function, oxidative-stress, inflammatory cytokines, and chemokines including monocyte chemoattractant protein-1 (MCP-1), lipid peroxidation, plasma proprotein convertase subtilisin/kexin type-9 (PCSK-9), and high-sensitivity C-reactive protein (hs-CRP). Moreover, the assessment of the hepatic mRNA expression of the key genes involved in cholesterol homeostasis depicted that SAC and SEC downregulated the PCSK-9 mRNA expression via targeting the expression of HNF-1α, a transcriptional activator of PCSK-9. On the other hand, the LDL-receptor (LDL-R) expression was upregulated through the activation of its transcriptional regulator sterol regulatory element binding protein-2 (SREBP-2). In addition, the activity and the mRNA expression of 3-hydroxy-3-methylglutaryl coenzyme-A reductases (HMG-R) and peroxisome proliferator-activated receptors (PPARs) were also improved by the treatment of SAC and SEC. We concluded that SAC and SEC can protect against MetS via improving the lipid and lipoprotein content, glycemic indices, hepatic function, targeting the inflammatory cascades, and oxidative imbalance, regulation of the mRNA expression of PCSK-9, LDL-R, SREBP-2, HNF-1α, PPARs, and inflammatory biomarkers.

A Comparative Study of LC-MS and FIA-(ESI)MS for Quantitation of S-Allyl-L-Cysteine in Aged Garlic Supplements.

The increasing consumption of food supplements demands the development of improved analytical methodologies to ensure their quality and authenticity. In this paper, two new approaches, liquid chromatography coupled to mass spectrometry (LC-MS) and flow injection analysis-(electrospray ionization) mass spectrometry (FIA-(ESI)MS), were optimized and validated for their application in the quantitative analysis of bioactive S-allyl-L-cysteine (SAC) in commercial aged garlic supplements (AGS). Although both methodologies were found to be useful for the sensitive and precise quantitation of SAC, the LC-MS approach allowed the differential determination of SAC and its bioactive diastereoisomer, S-1-propenyl-L-cysteine (S1PC), together with the identification of a number of organosulfur compounds typical of garlic. Mass fingerprints by FIA-(ESI)MS were proposed as an advantageous alternative to LC-MS analysis when the fast (4 min/sample) screening of AGS for their SAC content is intended, as in applications aimed at high-throughput quality control or standardization. Finally, the results gathered by the application of these two methodologies evidenced the highly variable composition of commercial AGS, as well as the identification of a number of potential composition frauds affecting their genuineness and benefits on health.

Characterization of a short-term processing technology of black garlic with low 5-HMF content

Black garlic, a deep-processed product of garlic, is widely favored in various Asian countries. Traditional black garlic production typically takes 60–90 days, resulting in low production efficiency and the formation of potentially harmful levels of 5-hydroxymethylfurfural (5-HMF). In this study, we implemented a novel production process capable of producing short-term black garlic (SBG) with significantly reduced levels of 5-HMF within just 8.6 h. A comparative analysis was conducted to investigate the quality disparities between SBG and commercially black garlic (CBG), encompassing assessments on color, physicochemical properties, characteristic components, in vitro antioxidant capacity, and volatile compounds. The results indicated that there were no significant differences in color between SBG and CBG. SBG exhibited higher levels of protein, reducing sugar, and polyphenols. 5-HMF, allicin, and S-allyl-l-cysteine (SAC) were selected as characteristic ingredients of black garlic. The concentration of 5-HMF in SBG was only 5.6% compared to CBG. SAC content in SBG was 2.3 times higher than in CBG and 11.3 times higher than in fresh garlic, while allicin was not detected in SBG samples. When comparing black garlic extracts at the same concentration, the antioxidant capacity of SBG was found to be stronger than that of CBG. Key flavor compounds identified in CBG included diallyl tetrasulfide, 5-methyl-2-furanmethanethiol, allyl methyl sulfide, dimethyl tetrasulfide, diallyl disulfide, diallyl sulfide, and dimethyl trisulfide. In contrast, key flavor compounds identified in SBG were 2-acetylthiophene, allyl methyl sulfide, 5-methyl-2-thiophenecarboxaldehyde, 2-acetylthiazole, dimethyl trisulfide, diallyl sulfide, and dimethyl disulfide. Compared to CBG samples, the aroma profile of SBGs exhibited a more pleasant scent with reduced pungency.

In-vitro antioxidant and anti-inflammatory potential along with p.o. pharmacokinetic profile of key bioactive phytocompounds of Snow Mountain Garlic: a comparative analysis vis-à-vis normal garlic.

Snow mountain garlic (SMG) is a trans-Himalayan medicinal plant used in the traditional medicine system for several ailments, including inflammatory arthritis. Research studies are insufficient to validate its folk medicinal applications. In the present study, the comparative abundance of its key bioactive phytocompounds, viz., S-allyl-L-cysteine (SAC), alliin, and S-methyl-L-cysteine (SMC) against normal garlic were assessed using the LC-MS/MS-MRM method. In addition, the study also explored the antioxidant and anti-inflammatory potency of crude extract of SMG and purified signature phytocompounds (i.e., SMC, SAC, and alliin) in comparison with normal garlic and dexamethasone in LPS-stimulated RAW264.7 macrophage cells. The LC-MS/MS-MRM study revealed significant differences among SMG and normal garlic, viz., alliin 22.8-fold higher in SMG, and SMC could be detected only in SMG. In the bioassays, SMG extract and purified signature phytocompounds significantly downregulated oxidative damage in activated macrophages, boosting endogenous antioxidants' activity. SMG extract-treated macrophages significantly suppressed NF-κB expression and related inflammatory indicators such as cytokines, COX-2, iNOS, and NO. Notably, the observed anti-inflammatory and antioxidant bioactivities of SMG extract were comparable to signature phytocompounds and dexamethasone. In addition, SAC being uniformly found in SMG and normal garlic, its comparative pharmacokinetics was studied to validate the pharmacodynamic superiority of SMG over normal garlic. Significantly higher plasma concentrations (Cmax), half-life (t1/2), and area under curve (AUC) of SAC following SMG extract administration than normal garlic validated the proposed hypothesis. Thus, the abundance of bioactive phytocompounds and their better pharmacokinetics in SMG extract might be underlying its medicinal merits over normal garlic.

S-Allyl-L-Cysteine Affects Cell Proliferation and Expression of H2S-Synthetizing Enzymes in MCF-7 and MDA-MB-231 Adenocarcinoma Cell Lines.

S-allyl-L-cysteine (SAC) is a sulfur compound present in fresh garlic. The reference literature describes its anticancer, antioxidant and neuroprotective effects. Breast cancer is infamously known as one of the most commonly diagnosed malignancies among women worldwide. Its morbidity and mortality make it reasonable to complete and expand knowledge on this cancer's characteristics. Hydrogen sulfide (H2S) and its naturally occurring donors are well-known investigation subjects for diverse therapeutic purposes. This study was conducted to investigate the SAC antiproliferative potential and effect on three enzymes involved in H2S metabolism: 3-mercaptopyruvate sulfurtransferase (MPST), cystathionine γ-lyase (CTH), and cystathionine β-synthase (CBS). We chose the in vitro cellular model of human breast adenocarcinomas: MCF-7 and MDA-MB-231. The expression of enzymes after 2, 4, 6, 8, and 24 h incubation with 2.24 mM, 3.37 mM, and 4.50 mM SAC concentrations was examined. The number of living cells was determined by the MTS assay. Changes in cellular plasma membrane integrity were measured by the LDH test. Expression changes at the protein level were analyzed using Western blot. A significant decrease in viable cells was registered for MCF-7 cells after all incubation times upon 4.50 mM SAC exposure, and after 6 and 24 h only in MDA-MB-231 upon 4.50 mM SAC. In both cell lines, the MPST gene expression significantly increased after the 24 h incubation with 4.50 mM SAC. S-allyl-L-cysteine had opposite effects on changes in CTH and CBS expression in both cell lines. In our research model, we confirmed the antiproliferative potential of SAC and concluded that our studies provided current information about the increase in MPST gene expression mediated by S-allyl-L-cysteine in the adenocarcinoma in vitro cellular model for the MCF-7 and MDA-MB-231 cell lines. Further investigation of this in vitro model can bring useful information regarding sulfur enzyme metabolism of breast adenocarcinoma and regulating its activity and expression (gene silencing) in anticancer therapy.

The Effect of S-Allyl L-Cysteine on Retinal Ischemia: The Contributions of MCP-1 and PKM2 in the Underlying Medicinal Properties.

Retinal ischemia plays a vital role in vision-threatening retinal ischemic disorders, such as diabetic retinopathy, age-related macular degeneration, glaucoma, etc. The aim of this study was to investigate the effects of S-allyl L-cysteine (SAC) and its associated therapeutic mechanism. Oxidative stress was induced by administration of 500 μM H2O2 for 24 h; SAC demonstrated a dose-dependent neuroprotective effect with significant cell viability effects at 100 μM, and it concurrently downregulated angiogenesis factor PKM2 and inflammatory biomarker MCP-1. In a Wistar rat model of high intraocular pressure (HIOP)-induced retinal ischemia and reperfusion (I/R), post-administration of 100 μM SAC counteracted the ischemic-associated reduction of ERG b-wave amplitude and fluorogold-labeled RGC reduction. This study supports that SAC could protect against retinal ischemia through its anti-oxidative, anti-angiogenic, anti-inflammatory, and neuroprotective properties.

The effect of S‐allyl L‐cysteine on retinal ischaemia: The role of MCP‐1 and PKM2

Aims/Purpose: Retinal ischaemia plays an important role in vision threatening retinal ischaemic disorders, such as central/branch RAO, nvAMD and normal tension glaucoma. The aim of this study was to determine the effects of S‐allyl L‐cysteine (SAC) and its underlying mechanism.Methods: Porcine RPE (pRPE) cells were extracted from adult pig eyes and maintained in DMEM containing 10% FBS, 100 U/mL penicillin and 100 mg/mL streptomycin in a humidified atmosphere of 95% air and 5% CO2 at 37°C. Cell viability was evaluated by via a haemocytometre under an optical microscope. H2O2 (500 μM) was added for 24 h to stimulate an oxidative stress and SAC or vehicle was added to some groups of the cultured pRPE cells 15 min before adding H2O2.Besides, high intraocular pressure (HIOP)‐induced retinal ischaemia was established by raising IOP to 120 mmHg for 60 min in a Wistar rat eye. This ischaemic insult is followed by 7 days of reperfusion. Whether and how ‘SAC’ is able to prevent retinal ischaemic injury were assessed by electrophysiology, fluorogold‐labeling and ELISA/western blot.Results: H2O2 (500 μM) stimulated free radical formation in cultured pRPE cells and significantly reduced the cell viability. This harmful effect was dose‐dependently (25/50 μM) and significantly (100 μM) ameliorated by pretreatment with SAC. Moreover, post‐administration of mega‐dose SAC (I/R + SAC 9 μg/kg/day ≈ 0.18 mg/0.2 Kg rat) counteracted the ischaemia‐induced decrease in the ERG b‐wave and reduction in the RGC cell number in the rat retina. This study supports an antioxidative role for SAC. Through the ELISA/western blot, SAC (100 μM) appears to protect against H2O2‐induced oxidative stress via the downregulation of ischaemia‐related factor PKM2 and inflammatory biomarker MCP‐1.Conclusions: SAC might protect against retinal ischaemia by antioxidation, anti‐ischaemia and anti‐inflammation (downregulating PKM2 and MCP‐1).

Preparation of novel S-allyl cysteine chitosan based nanoparticles for use in ischemic brain treatment.

Objective: To enhance the brain bioavailability of S-allyl-l-cysteine (SC) by developing novel S-allyl-l-cysteine chitosan nanoparticles (SC CS NPs) and examining the quantity of SC by developing a novel method of ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) in ischemic rat brain treatment. Methods: The ionotropic gelation method was used to develop S-allyl cysteine-loaded CS NPs. The 4-factor, 5-level central composite design was optimized to determine the effect of independent variables, i.e., particle size, polydispersity index (PDI), zeta potential, EE, and loading capacity, together with their characterization, followed by drug release and intranasal permeation to enhance the brain bioavailability and examination of their neurobehavioral and biochemical parameters with their histopathological examination. Results: SC CS NPs were optimized at the particle size of 93.21 ± 3.31 nm (PDI: 0.317 ± 0.003), zeta potential of 44.4 ± 2.93, and drug loading of 41.23 ± 1.97% with an entrapment efficiency of 82.61 ± 4.93% having sustain and controlled release (79.92 ± 3.86%) with great permeation (>80.0%) of SC. SC showed the retention time of 1.021 min and 162.50/73.05 m/z. SC showed good linearity in the range of 5.0-1300.0 ng mL-1, % inter-and-intraday accuracy of 96.00-99.06% and CV of 4.38-4.38%. We observed significant results, i.e., p < 0.001 for improved (AUC)0-24 and Cmax delivered via i.v. and i.n. dose. We also observed the highly significantly observations of SC CS NPs (i.n.) based on their treatment results for the biochemical, neurobehavioral, and histopathological examination in the developed ischemic MCAO brain rat model. Conclusion: The excellent significant role of mucoadhesive CS NPs of SC was proven based on the enhancement in the brain bioavailability of SC via i.n. delivery in rats and easy targeting of the brain for ischemic brain treatment followed by an improvement in neuroprotection based on a very small dose of SC.

Naturally occurring organosulfur compounds effectively inhibits PCSK-9 activity and restrict PCSK-9-LDL-receptor interaction via in-silico and in-vitro approach

The present study intended to divulge the potential role of garlic-derived organosulfur compounds (OSCs) in targeting PCSK-9 and averting its interaction with the EGF-A portion of LDL-R via in-vitro and in-silico analysis. Our in-silico screening data showed that 3-(Propylsulfinyl)-L-alanine (PSA), S-Ethyl-L-cysteine (SEC), alliin, and S-Allyl-L-cysteine (SAC) exhibited higher binding energy (−7.05, −7.00, −6.65, and −6.31 Kcal/mol, respectively) against PCSK-9, among other selected OSCs. Further, the protein-protein interaction study of PCSK-9-OSCs-complex with EGF-A demonstrated a similar binding pattern with E-total values ranging from −430.01 to −405.6 Kcal/mol. These results were further validated via in-vitro analysis which showed that SEC, SAC, and diallyl trisulphide (DAT) exhibited the lowest IC50 values of 4.70, 5.26, and 5.29 µg/mL, respectively. In conclusion, the presented data illustrated that SEC, SAC, and DAT were the best inhibitors of PCSK-9 activity and may have the potential to improve the LDL-R function and lower the circulatory LDL-C level.

Optimization of complex coacervation parameters for the production of encapsulated black garlic using response surface methodology.

The purpose of this study was to optimize black garlic encapsulation parameters (core/coating ratio, extract concentration, and coacervate/maltodextrin [MD] ratio) using central composite design of the response surface methodology based on encapsulation efficiency (EE) (%). The optimum parameters were determined as 4.0 for the coating material/core ratio, 50% for the extract concentration, and 6.0 for the MD/coacervate ratio depending on the EE (%). The antioxidant activity values were determined as 101 and 134µmol Trolox/100g dry weight (DW) for the 2,2-diphenyl-1-picrylhydrazyl and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) methods, respectively, whereas the total phenolic content was 49mg gallic acid equivalent/100gDW for the encapsulated black garlic samples. S-Allyl-l-cysteine (SAC), γ-l-glutamyl-SAC (GSAC), γ-l-glutamyl-(S)-trans-1-propenyl-l-cysteine, and allicin were the organosulfur (OS) compounds determined in the samples. The SAC concentration of the encapsulated black garlic samples was determined as 22.36mg/g, whereas the GSAC content was found at a lower concentration (0.33mg/g) compared to SAC. The allicin content was quantified to be 0.31mg/g. The encapsulated samples were also characterized by scanning electron microscopy (SEM) and Fourier transform infrared (FT-IR) spectroscopy. The FT-IR analysis revealed specific functional groups, including hydroxyl, carbonyl, and glycosidic linkage. The interaction between lentil protein isolate and pectin was strong enough to encourage capsule formation as visualized in the SEM images. This study shows the potential of black garlic coacervates as a functional ingredient for the food industry due to their stability, solubility, and preservation of OS and antioxidant compounds.

Simultaneous microwave-assisted extraction of bioactive compounds from aged garlic

In this work, the simultaneous extraction of bioactives (organosulfur compounds, such as S-allyl-L-cysteine (SAC), carbohydrates, such as neokestose and neonystose, and total phenolic compounds) from aged garlic has been optimized for the first time to obtain multifunctional extracts for further application as food ingredients. Analytical methods using liquid chromatography coupled to mass spectrometry (HPLC-MS) and by hydrophilic interaction liquid chromatography with evaporative light scattering detection (HILIC-ELSD) were also previously optimized. High sensitivity (limits of detection between 0.013 and 0.77 µg mL−1) and appropriate repeatability (< 12%) and accuracy (> 92%) for the analysis of bioactives were achieved. After selecting water as the extraction solvent and microwave-assisted extraction (MAE) as the most efficient technique, operation conditions were optimized using a Box-Behnken experimental design (60 min; 120 °C; 0.05 g mL−1; 1 cycle) to maximize the content of bioactives from different aged garlic samples. Regarding organosulfur compounds, only SAC (traces-2.32 mg g−1 dry sample) and cycloalliin (1.23–3.01 mg g−1 dry sample) were detected in all samples, while amino acids such as arginine (0.24–3.45 mg g−1 dry sample) and proline (0.43–3.91 mg g−1 dry sample) were, in general, the most abundant. Bioactive carbohydrates (from trisaccharides to nonasaccharides) were only detected in fresh garlic and aged garlic processed under mild conditions, whereas all garlic extracts showed antioxidant activity. The developed MAE methodology is shown as a successful alternative to other procedures for the simultaneous extraction of aged garlic bioactives intended by the food and nutraceutical industries, among others.

Bioavailability of Organosulfur Compounds after the Ingestion of Black Garlic by Healthy Humans.

The consumption of black garlic has been related to a decreased risk of many human diseases due to the presence of phytochemicals such as organosulfur compounds (OSCs). However, information on the metabolization of these compounds in humans is limited. By means of ultra-high-performance liquid chromatography coupled with high-resolution mass spectrometry (UHPLC-HRMS), this study aims to determine the OSCs and their metabolites excreted in urine 24 h after an acute intake of 20 g of black garlic by healthy humans. Thirty-three OSCs were identified and quantified, methiin (17,954 ± 6040 nmol), isoalliin (15,001 ± 9241 nmol), S-(2-carboxypropyl)-L-cysteine (8804 ± 7220 nmol) and S-propyl-L-cysteine (deoxypropiin) (7035 ± 1392 nmol) being the main ones. Also detected were the metabolites N-acetyl-S-allyl-L-cysteine (NASAC), N-acetyl-S-allyl-L-cysteine sulfoxide (NASACS) and N-acetyl-S-(2-carboxypropyl)-L-cysteine (NACPC), derived from S-allyl-L-cysteine (SAC), alliin and S-(2-carboxypropyl)-L-cysteine, respectively. These compounds are potentially N-acetylated in the liver and kidney. The total excretion of OSCs 24 h after the ingestion of black garlic was 64,312 ± 26,584 nmol. A tentative metabolic pathway has been proposed for OSCs in humans.

S-ALLYL-L-CYSTEINE-INDUCED ANTI-INFLAMMATORY AND ANTI-APOPTOTIC EFFECTS IN CHONDROCYTES IS ASSOCIATED WITH SUPPRESSION OF THE MITOCHONDRIAL INFLAMMATION PATHWAY

One major aspects to consider while dealing with osteoarthritis is oxidative stress. This deleterious oxidative stress is responsible for the increased production of Reactive Oxygen Species and triggers several inflammatory pathways, including Mitochondrial Inflammation Pathway (MIP), which leads the cell to apoptosis. Chondrocytes, under oxidative stress, are unable to synthesize cartilage efficiently. S-Allyl-L-Cysteine (SAC) is known to be a potent natural, water-soluble antioxidant derived from garlic whose antioxidant properties have been evaluated in several diseases at the molecular level; other than osteoarthritis. Herein, we investigated the potential of S-Allyl-L-Cysteine (SAC) preconditioning of chondrocytes against oxidative stress-mediated mitochondrial inflammation. SAC priming alleviated oxidative stress-induced injuries by significantly improved cell viability, morphology and activated cell migration. In addition, decreased lactate dehydrogenease, increased superoxide dismutase release and retention of glycosaminoglycans were observed.SAC preconditioning ameliorated the injurious effects of oxidative stress as revealed by significant downregulation in gene expression of hypoxia-induciblefactor 1α (Hif-1α), Xanthine Oxidase (XO), Caspase-9 (Casp-9), Caspase-3(Casp-3), Interleukin 1 beta (IL-1β) and inducible nitric oxide synthase (iNOS). These findings suggest that SAC preconditioning might enhance the antioxidant and anti-inflammatory efficacy of chondrocytes by regulating the MIP pathway and improving cellular responses.

Encapsulated black garlic: Comparison with black garlic extract in terms of physicochemical properties, biological activities, production efficiency and storage stability

In this paper, the convenience of encapsulation technique was investigated in terms of characterization of capsule, changing the volatile compounds and preservation biological activity throughout six months for black garlic (BG) by comparison with the aqueous extract which prepared from BG (stored at 4 °C) each analysis period. Besides, the genotoxic and antigenotoxic aspects of aqueous black garlic extract (BGE) and black garlic capsule (BGC) developed as a novel promising product were also researched. The S-allyl-L-cysteine (SAC) content, which is mostly known to be responsible for biological activity, was preserved in BGC during six months. The variety of volatile compound of BGC (2 compounds) was significantly reduced compared to BG (17 compounds) and BGE (7 compounds), which indicates that the extraction and encapsulation traps the undesirable odor. A significant decrease was observed for total phenolic content and antioxidant capacity (performed with DPPH and CUPric Reducing Antioxidant Capacity (CUPRAC)) of BGC with storage like BGE. In contrast, the values obtained with the ABTS method remained constant with storage. The antimicrobial activity (carried out on five Gram-positive bacteria, three Gram-negative bacteria and three yeasts) of BGC was preserved better than BGE. The treatment concentrations (50, 100, 200 μg/mL) of both BGE and BGC did not have a genotoxic effect and moreover they have antigenotoxic effects against mitomycin C on human lymphocyte cells. All these results implied that BGC can be good a candidate to increase the consumption of BG providing health benefits and functional supplement for food.

Identification of tryptophanase from Escherichia coli for the synthesis of S-allyl-l-cysteine and related S-substituted cysteine derivatives

A wide variety of S-substituted cysteine derivatives occur in plant metabolites. For example, S-allyl-l-cysteine (SAC), mainly contained in garlic, gathers huge interest because of its favorable bioactivities for human health. However, conventional methods for preparing SAC suffer from several drawbacks with regard to efficiency and toxicity, which highlights the need for improved processes for SAC synthesis. This study aims to develop a novel bioprocess to produce SAC by microbial enzymes from easily available substrates. We found that Escherichia coli had the ability to synthesize SAC from allyl mercaptan, pyruvic acid, and ammonium sulfate. An enzyme purification through 3-step column chromatography, followed by determination of the N-terminal amino acid sequence revealed that tryptophanase (TnaA) was the enzyme responsible for SAC formation. Although the enzyme catalyzed the reversible reaction for synthesizing and degrading SAC, the degradation proceeded significantly faster than the synthesis. Interestingly, TnaA catalyzed the synthesis of a wide range of S-substituted cysteines with alkyl chains or aromatic rings, some of which are present in Allium and Petiveria plants. Our results showed a novel substrate specificity of TnaA toward various S-substituted cysteine. TnaA is a promising biocatalyst for developing a new process to supply various valuable S-substituted cysteine derivatives for medicinal and health-promoting applications.

Antioxidant Interactions between S-allyl-L-cysteine and Polyphenols Using Interaction Index and Isobolographic Analysis.

This work aims to study the antioxidant interactions between S-allyl-L-cysteine (SAC) and six natural polyphenols (quercetin, caffeic acid, sinapic acid, catechin, ferulic acid, and 3,4-dihydroxybenzoic acid) through the measurement of free-radical-scavenging activity of 1,1-diphenyl- 2-picryl-hydrazyl (DPPH), the radical-cation-scavenging activity of 2,2-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS), and reducing power. Among the six natural polyphenols, caffeic acid showed the strongest synergistic effect with SAC according to DPPH and reducing power assays. Further investigations based on the results of interaction index and isobologram analysis showed that the antioxidant activity (DPPH, ABTS, and reducing power) of the combination of caffeic acid with SAC presented an increase with the raising of their individual concentrations in their mixture and along with a dose–response manner. The best synergistic effect between caffeic acid and SAC based on DPPH, ABTS, and reducing power assays were observed at the ratio of 1:20, 1:35, and 1:70, respectively. The excellent synergic antioxidant activity of the combination of caffeic acid with SAC in our study suggests SAC has a more broad and effective application prospects in food field.

Cell proliferation effects of S-allyl-L-cysteine are associated with phosphorylation of janus kinase 2, insulin-like growth factor type-I receptor tyrosine kinase, and extracellular signal-regulated kinase 2 in primary cultures of adult rat hepatocytes

The cell proliferation effect of S-allyl-L-cysteine (SAC) and its mechanisms were examined in primary cultures of adult rat hepatocytes. In serum-free cultivation, SAC (10−6 M)-stimulated hepatocytes showed significant proliferation compared to control at 5-h culture; the effect was dependent on the culture time and the dose of SAC (EC50 value 8.58 × 10−8 M). In addition, SAC-stimulated hepatocytes significantly increased mRNA expression levels of c-Myc and c-Fos at 1 h and cyclin B1 at 3.5 and 4 h, respectively. In contrast, alliin and allicin, structural analogs of SAC, did not show these effects observed with SAC. The SAC-induced hepatocyte proliferation effects were completely suppressed by monoclonal antibodies against growth hormone receptor and insulin-like growth factor type-I (IGF-I) receptor, respectively. Furthermore, the Janus kinase 2 (JAK2) inhibitor TG101209, phospholipase C (PLC) inhibitor U-73122, IGF-I receptor tyrosine kinase (RTK) inhibitor AG538, PI3 kinase inhibitor LY294002, MEK inhibitor PD98059, and mTOR inhibitor rapamycin completely suppressed the SAC-induced hepatocyte proliferation. JAK2 (p125 kDa) phosphorylation in cultured hepatocytes peaked 5 min after SAC stimulation. SAC-induced IGF-I RTK (p95 kDa) and ERK2 (p42 kDa) phosphorylation had slower rises than JAK2, peaking at 20 and 30 min, respectively. These results indicate that SAC promoted cell proliferation by growth hormone receptor/JAK2/PLC pathway activation followed by activation of the IGF-I RTK/PI3K/ERK2/mTOR pathway in primary cultures of adult rat hepatocytes.